Effect of Tooth Loss on Cognitive Function among Older Adults in Singapore.

Evidence suggests a longitudinal association between tooth loss and cognitive function. However, the temporality of this association is not well understood. We investigated the effect of several emulated tooth loss prevention scenarios on cognitive function. We used data from 3 waves (baseline: 2009, second wave: 2011-2012, and third wave: 2015) of the Panel on Health and Ageing of Singaporean Elderly (PHASE). PHASE targeted older adults, aged ≥60 y, in Singapore. Number of teeth was used as a time-varying exposure (baseline, second wave). Cognitive function (Short Portable Mental Status Questionnaire score) in the third wave was assessed as the outcome. Multiple time-invariant (baseline) and time-varying (baseline and second wave) covariates were included. The "longitudinal modified treatment policy approach" combined with targeted minimum loss-based estimation was used to define and estimate additive effects of emulated tooth loss prevention scenarios. Emulated scenarios were the following: what if edentate people retained 1 to 4 teeth (scenario 1), what if those with <5 teeth retained 5 to 9 teeth (scenario 2), what if those with <10 teeth retained 10 to 19 teeth (scenario 3), and what if everyone retained ≥20 teeth (scenario 4)? A total of 1,516 participants, excluding those with severe cognitive impairment, were included (male: 41.6%). The mean age at baseline was 70.6 y (SD = 7.1). The mean SPMSQ score at baseline was 2.06 (SD = 0.02) for edentulous, 1.55 (SD = 0.04) for 1 to 4 teeth, 1.61 (SD = 0.03) for 5 to 9 teeth, 1.73 (SD = 0.02) for 10 to 19 teeth, and 1.71 (SD = 0.02) for ≥20 teeth. Additive effect of hypothetical intervention gradually increased with intensity of prevention from scenario 1 to scenario 4 (scenario 1: -0.02 [95% CI, -0.08 to 0.04], scenario 2: -0.05 [95% CI, -0.11 to -0.00], scenario 3: -0.07 [95% CI, -0.14 to -0.00], scenario 4: -0.15 [95% CI, -0.23 to -0.06]). Emulated tooth loss prevention interventions were associated with better cognitive function score. Therefore, preventing tooth loss could potentially benefit maintenance of cognitive function among older adults.

Weight Loss Mediated the Relationship between Tooth Loss and Mortality Risk.

Tooth loss is a risk factor for increased mortality; however, the underlying mechanism remains unclear. This study aimed to evaluate the mediating effect of weight change on the relationship between tooth loss and mortality risk. This was a 10-y follow-up prospective cohort study using the data from the Japan Gerontological Evaluation Study (JAGES). The participants were independent older adults aged ≥65 y at baseline and were followed up from 2010 to 2020. The incidence of death in 2013 and 2020, incidence of >5% weight loss/gain in 2010 and 2013, and the number of remaining teeth in 2010 were used as the outcome, mediator, and explanatory variables, respectively. We conducted causal mediation analysis by fitting the Cox proportional hazard model, including possible confounders. Hazard ratios (HRs) and 95% confidence intervals (95% CIs) of the total effect (TE), natural indirect effect (NIE), and proportion mediated (PM) were estimated. Among the 34,510 participants, the mean age was 72.6 (SD = 5.4) y, and 47.6% were men. From 2013 to 2020, 14.0% of the participants (n = 4,825) died, 60.5% (n = 20,871) had 0 to 19 remaining teeth, and 17.2% (n = 5,927) and 8.4% (n = 2,907) experienced >5% weight loss and gain, respectively. The mortality rate was 0.016 per person-year among those with ≥20 remaining teeth and 0.027 per person-year among those with 0 to 19 remaining teeth. Weight loss of >5% significantly mediated the association between tooth loss and higher mortality risk (TE: HR, 1.28 [95% CI, 1.16 to 1.40]; NIE: HR, 1.03 [95% CI, 1.02 to 1.04]; PM, 13.1%); however, we observed a slight mediating effect for >5% weight gain (NIE: HR, 1.003 [95%CI, 1.0001 to 1.01]; PM, 1.3%). The present study suggests that a clinically meaningful level of weight loss mediated the association between tooth loss and increased risk of mortality among independent older adults.

Oral Status and Dementia Onset: Mediation of Nutritional and Social Factors.

Some modifiable risk factors for dementia are closely related to oral health. Although eating and speaking abilities are fundamental oral functions, limited studies have focused on the effect of malnutrition and lack of social interaction between oral health and dementia. We investigated the mediating effects of nutritional and social factors on the association between the number of teeth and the incidence of dementia. This 6-y cohort study used data from the Japan Gerontological Evaluation Study targeting older adults aged 65 y and above. The number of teeth (exposure) and covariates in 2010 (baseline survey), mediators (weight loss, vegetable and fruit intake, homeboundness, social network) in 2013, and the onset of dementia (outcome) between 2013 and 2016 were obtained. The Karlson-Holm-Breen mediation method was applied. A total of 35,744 participants were included (54.0% women). The mean age at baseline was 73.1 ± 5.5 y for men and 73.2 ± 5.5 y for women. A total of 1,776 participants (5.0%) had dementia during the follow-up period. There was a significant total effect of the number of teeth on the onset of dementia (hazard ratio, 1.14; 95% CI, 1.01-1.28). Controlling for nutritional and social mediators, the effect of the number of teeth was reduced to 1.10 (95% CI, 0.98-1.25), leaving an indirect effect of 1.03 (95% CI, 1.02-1.04). In the sex-stratified analysis, the proportion mediated by weight loss was 6.35% for men and 4.07% for women. The proportions mediated by vegetable and fruit intake and homeboundness were 4.44% and 4.83% for men and 8.45% and 0.93% for women, respectively. Furthermore, the proportion mediated by social networks was 13.79% for men and 4.00% for women. Tooth loss was associated with the onset of dementia. Nutritional and social factors partially mediated this association.

Effect of Copayment on Dental Visits: A Regression Discontinuity Analysis.

Despite their prevalence and burdens, oral diseases are neglected in universal health coverage. In Japan, a 30% copayment (out of pocket) by the user and a 70% contribution by Japan's universal health insurance (JUHI) are required for dental and medical services. From the age of 70 y, an additional 10% is offered by JUHI (copayment, 20%; JUHI, 80%). This study aimed to investigate the effect of cost on dental service use among older adults under the current JUHI system. A regression discontinuity quasi-experimental method was used to investigate the causal effect of the JUHI discount policy on dental visits based on cross-sectional data. Data were derived from the 2016 Japan Gerontological Evaluation Study. This analysis contained 7,161 participants who used JUHI, were aged 68 to 73 y, and responded to questions regarding past dental visits. Analyses were controlled for age, sex, number of teeth, and equalized household income. Mean ± SD age was 72.1 ± 0.79 y for the discount-eligible group and 68.9 ± 0.78 y for the noneligible group. During the past 12 mo, significantly more discount-eligible participants had visited dental services than noneligible participants (66.0% vs. 62.1% for treatment visits, 57.7% vs. 53.1% for checkups). After controlling for covariates, the effect of discount eligibility was significant on dental treatment visits (odds ratio [OR], 1.36; 95% CI, 1.32 to 1.40) and dental checkups (OR, 1.49; 95% CI, 1.44 to 1.54) in the regression discontinuity analysis. Similar findings were observed in triangular kernel-weighted models (OR, 1.38 [95% CI, 1.34 to 1.44]; OR, 1.52 [95% CI, 1.47 to 1.56], respectively). JUHI copayment discount policy increases oral health service utilization among older Japanese. The price elasticity for dental checkup visits appears to be higher than for dental treatment visits. Hence, reforming the universal health coverage system to improve the affordability of relatively inexpensive preventive care could increase dental service utilization in Japan.

Genomic structure of swine taste receptor family 1 member 3, TAS1R3, and its expression in tissues.

Taste receptor family 1 member 3, TAS1R3, is shown to be involved in sweet and umami tastes in mouse, and the nucleotide sequence of the gene has been reported in rat, gorilla, and human. Pigs are frequently used as models for human diseases, and are also considered to be source animals for xenotransplantation to humans due to their anatomical and physiological similarities to humans. Therefore, in the present study, the genomic structure of the swine TAS1R3 gene was determined, and TAS1R3 expression was studied in various swine tissues. The gene was shown to reside on swine chromosome 6q22-->q23, from which three types of mRNAs were generated: 3,752 bp derived from six exons in tongue, 3,704 bp from six exons and 3,630 bp from seven exons in testis. The 6 exons/5 introns were structurally similar to those of humans and mice, but the 7 exons/6 introns structure of TAS1R3 was first observed in swine. High expressions of TAS1R3 were revealed in tongue, kidney, and testis by real-time PCR. The expression profile of the tissues except for kidney was similar to that of mouse. When in situ hybridization using an RNA probe for TAS1R3 was performed on swine tongue and testis tissues, TAS1R3 expressions were revealed in tongue circumvallate papillae, fungiform papillae, mucosal epithelium, follicular B lymphocytes, lymphocytes in submucosal tissues of lingual tonsil, and spermatogenic cells. Using peripheral mature B lymphocytes, the expression of TAS1R3 in B lymphocytes was further confirmed by real-time PCR and sequencing of the real-time PCR product.

Assignment of 117 genes from HSA5 to the porcine IMpRH map and generation of a dense human-pig comparative map.

Twenty-two and eight significant quantitative trait loci for economically important traits have been located on porcine chromosomes (SSC) 2q and SSC16 respectively, both of which have been shown to correspond to human chromosome 5 (HSA5) by chromosome painting. To provide a comprehensive comparative map for efficient selection of candidate genes, we assigned 117 genes from HSA5 using a porcine radiation hybrid (IMpRH) panel. Sixty-six genes were assigned to SSC2 and 48 to SSC16. One gene was suggested to link to SSC2 markers and another to SSC6. One gene did not link to any gene, expressed sequence tag or marker in the map, including those in the present investigation. This study demonstrated the following: (1) SSC2q21-q28 corresponds to the region ranging from 74.0 to 148.2 Mb on HSA5q13-q32 and the region from 176.0 to 179.3 Mb on HSA5q35; (2) SSC16 corresponds to the region from 1.4 to 68.7 Mb on HSA5p-q13 and to the region from 150.4 to 169.1 Mb on HSA5q32-q35 and (3) the conserved synteny between HSA5 and SSC2q21-q28 is interrupted by at least two sites and the synteny between HSA5 and SSC16 is also interrupted by at least two sites.

Assignment of 204 genes localized on HSA17 to a porcine RH (IMpRH) map to generate a dense comparative map between pig and human/mouse.

Bi- and uni-directional chromosome painting (ZOO-FISH) and gene mapping have revealed correspondences between human chromosome (HSA) 17 and porcine chromosome (SSC) 12 harboring economically important quantitative trait loci. In the present study, we have assigned 204 genes localized on HSA17 to SSC12 to generate a comprehensive comparative map between HSA17 and SSC12. Two hundred fifty-five primer pairs were designed using porcine sequences orthologous with human genes. Of the 255 primer pairs, 208 (81.6%) were used to assign the corresponding genes to porcine chromosomes using the INRA-Minnesota 7000-rad porcine x Chinese hamster whole genome radiation hybrid (IMpRH) panel. Two hundred three genes were integrated into the SSC12 IMpRH linkage maps; and one gene, PPARBP, was found to link to THRA1 located in SSC12 but not incorporated into the linkage maps. Three genes (GIT1, SLC25A11, and HT008) were suggested to link to SSC12 markers, and the remaining gene (RPL26) did not link to any genes/expressed sequence tags/markers registered, including those in the present study. A comparison of the gene orders among SSC12, HSA17, and mouse chromosome 11 indicates that intra-chromosomal rearrangements occurred frequently in this ancestral mammalian chromosome during speciation.

Assignment of 101 genes localized in HSA10 to a swine RH (IMpRH) map to generate a dense human-swine comparative map.

Economically important traits such as growth and backfat in pigs have been shown to be influenced by genes in swine chromosome (SSC) 10q12-->qter corresponding to human chromosome (HSA) 10p. However, since gene information in the swine chromosomal region was limited, we attempted to generate a dense comparative map between SSC10 and HSA10 by mapping the 115 genes of HSA10 to a swine RH map (IMpRH map). In the mapping ten genes were assigned to SSC10, 88 to SSC14, and one to SSC3. One gene was suggested to link to SSC3, and another to SSC9. The correspondences between HSA10 and SSC10 and between HSA10 and SSC14 were essentially consistent with the observations obtained from bi/uni-directional chromosome painting or other results. This study further indicated that a large number of intrachromosomal rearrangements occurred in the synteny-conserved regions following species separation.

A dense comparative gene map between human chromosome 19q13.3-->q13.4 and a homologous segment of swine chromosome 6.

The human chromosome (HSA)19q region has been shown to correspond to swine chromosome (SSC) 6q11-->q21 by bi-directional chromosomal painting and gene mapping. However, since the precise correspondence has not been determined, 26 genes localized in HSA19q13.3-->q13.4 were assigned to the SSC6 region mainly by radiation hybrid (RH) mapping, and additionally, by somatic cell hybrid panel (SCHP) mapping, and fluorescent in situ hybridization (FISH). Out of the 26 genes, 24 were assigned to a swine RH map with LOD scores greater than 6 (threshold of significance). The most likely order of the 24 genes along SSC6 was calculated by CarthaGene, revealing that the order is essentially the same as that in HSA19q13.3-->q13.4. For AURKC and RPS5 giving LOD scores not greater than 6, SCHP mapping and FISH were additionally performed; SCHP mapping assigned AURKC and RPS5 to SSC6q22-->q23 and SSC6q21, respectively, which is consistent with the observation of FISH. Consequently, all the genes (26 genes) examined in the present study were shown to localize in SSC6q12-->q23, and the order of the genes along the chromosomes was shown to be essentially the same in swine and human, though several intrachromosomal rearrangements were observed between the species.

Radiation hybrid map assignments of 11 ESTs obtained from a 28-day-old swine embryo cDNA library to the IMpRH map.

In order to improve the map resolution and to locate more genes on the porcine radiation hybrid map, expressed sequence tags (ESTs) were isolated from a 28-day-old normal pig embryo cDNA library. The ESTs were sequenced from the 5'-end and similarities were checked with sequences registered in the NCBI DNA database (http://www.ncbi.nlm.nih.gov/blast/). The ESTs sequences which have high identity scores (>80%) against human genes or ESTs were further sequenced from the 3' untranslated region. The ESTs which were sequenced successfully were used to design primers for PCR analysis of the radiation hybrid panel. Eleven ESTs were physically mapped to porcine chromosomes 2, 4, 8, 10, 13, 14 and X. The localizations are in agreement with the comparative mapping data between human and pig. The results will provide unique information to the comparative map of human and pig.

Conservation of the syntenies between porcine chromosome 7 and human chromosomes 6, 14 and 15 demonstrated by radiation hybrid mapping and linkage analysis.

Comparative mapping studies facilitate the identification of genes located in quantitative trait locus (QTL) regions in domestic animals by utilizing information from the human genome. Radiation hybrid (RH) mapping is effective for this purpose because of its high resolution in ordered gene mapping on chromosomes. We constructed an RH map of pig chromosome 7, by adding 23 markers associated with genes. This RH map clearly demonstrated the mosaic of homology between pig chromosome 7 (SSC7) and human chromosomes 6, 14 and 15 at a 'gene' level, and was confirmed by linkage analysis. Clarification of the homology of SSC7 to human chromosomes will contribute to the elucidation of the gene(s) responsible for QTL detected on this chromosome.

Construction of a high-resolution comparative gene map between swine chromosome region 6q11-->q21 and human chromosome 19 q-arm by RH mapping of 51 genes.

A comprehensive and comparative map was constructed for the porcine chromosome (SSC) 6q11-->q21 region, where the gene(s) responsible for the maldevelopment of embryos are localized using swine populations of the National Institute of Animal Industry, Japan (NIAI). Since the chromosomal region corresponds to a region of human chromosome (HSA) 19q13.1-->q13.3 based on bi-directional chromosome painting, primer pairs were designed from porcine cDNA sequences identified, on a sequence comparison basis, as being transcripts from genes orthologous to those in the HSA region. Fifty-one genes were successfully assigned to a swine radiation hybrid (RH) map with LOD scores greater than 6. ERF and PSMD8 genes were assigned to SSC4 and SSC1, respectively. The remaining 49 genes were assigned to SSC6, demonstrating that the synteny between the SSC6 and HSA19 chromosomal regions is essentially conserved, therefore confirming, the results of bi-directional chromosome painting. However, when examined precisely, rearrangements have apparently occurred within the region of conserved synteny. For the ERF and PSMD8 genes assigned to SSCs other than SSC6, additional mapping using somatic cell hybrid (SCH) panels was performed to confirm the results of RH-mapping.

Construction of a dense comparative map between human chromosome 1p36-->p35 and swine chromosome 6 by using human sequence-tagged sites.

Construction of a comprehensive comparative map between swine and human chromosomes is a prerequisite, in order to select candidate swine genes for traits from the human genome database as well as to understand the evolutionary process of the two species. The present study attempted to use 910 sequence-tagged sites (STSs) localized in human chromosome (HSA) 1p36-->p35 (35 Mbp) for radiation hybrid (RH) mapping to swine chromosomes (SSCs). Out of the 910 STSs subjected to amplification of swine orthologues, primer pairs for 13 STSs were found to amplify the respective orthologues and the STSs were assigned to SSCs. Eleven STSs were assigned to SSC6 in the same order as that in HSA1: SSC6cen-(SHGC-150)-(A006H31)-(X82877)-(A007E03)-(IB404)-(stGDB:371372)-(stSG31658)-(A009Q18)-(stSG14201/A009C01)-(H08335)-qter. One of the remaining two STSs, WI-20819, was assigned to SSCX, and the other, R91D18R, was not linked to any first-generation markers of the IMpRH map with a lod score greater than 3.

Genomic organization and assignment of the interleukin 7 gene (IL7) to porcine chromosome 4q11-->q13 by FISH and by analysis of radiation hybrid panels.

Interleukin 7 (IL7) is a cytokine that has many immunological functions, including regulation of hematopoiesis and peripheral lymphocytes. cDNA and a genomic DNA segment containing the porcine IL7 gene were isolated and sequenced, showing that porcine IL7 consists of 176 amino acids and that its gene spans over about 13 kb of genomic DNA. Porcine IL7 has 85% and 73% homology with human IL7 in terms of the nucleotide and amino acid sequences, respectively. Whereas the murine IL7 gene does not have an exon corresponding to human exon 5 (Lupton et al., 1990), the porcine IL7 gene was found to contain the same exon-intron structure as the human gene. These findings, together with the upstream structure of the cDNA elucidated in the present study, indicate that the relationship between swine and human IL7 is closer than that between mouse and human IL7. The IL7 gene was mapped to swine chromosome 4q11-->q13 by fluorescence in situ hybridization and, using a radiation hybrid panel, was localized between microsatellite markers Sw1336 and Sw1073 on the same chromosome.

Assignment of T cell receptor (TCR) alpha-chain gene (A), beta-chain gene (B), gamma-chain gene (G), and delta-chain gene (D) loci on swine chromosomes by in situ hybridization and radiation hybrid mapping.

Using the cDNA sequence of porcine T-cell receptor (TCR) alpha-, beta-, gamma-, and delta-chain genes, we screened a porcine BAC library to isolate clones containing these genes. The isolated BAC clones were confirmed to carry these TCR genes by partial nucleotide sequencing. Among the clones obtained in the present screening, two clones carried both the TCR alpha-chain gene (TRA) and the TCR delta-chain gene (TRD) while one clone each carried only the sequence of either TRA or TRD. This observation demonstrated that TRA and TRD are localized in close proximity on a swine chromosome. Also two clones contained the sequence of the TCR beta-chain gene (TRB), and two clones contained the sequence of TCR gamma-chain gene (TRG). Fluorescence in situ hybridization using the above BAC clones as probes revealed that TRA and TRD, TRB, and TRG loci reside on swine chromosomes 7q15.3-->q21, 18q11.3-->q12, and 9q21-->q22, respectively. The chromosome positions of TRA and TRB are consistent with those determined by somatic cell hybrid analysis (Rettenberger et al., 1996). In addition, RH mapping of these genes was performed using the INRA-University of Minnesota RH panel DNA. The result confirmed the position of TRA and TRB reported earlier (http://imprh.toulouse.inra.fr/), and further demonstrated that TRG was located 11 cR away from genetic marker SW989 toward the marker S0019.

Assignment of 64 genes expressed in 28-day-old pig embryo to radiation hybrid map.

A swine resource family was constructed at the National Institute of Animal Industry, Japan, in order to determine the genetic regions responsible for economically important traits, including fetus development. To identify genes expressed in the early stage of embryo development, we cataloged and mapped genes expressed in a 28-day-old normal pig embryo. In this effort, we have mapped 64 genes, which have map information in human genome onto a swine radiation hybrid (RH) map, IMpRH. These mappings provided additional chromosomal homologies between swine and human to improve the comparative map between the two species. The distribution of the genes assigned to swine chromosomes are as follows: 9 genes were assigned on SSC6; 6 genes each assigned on SSC5 and SSC14; 5 genes each assigned on SSC3, SSC4, and SSC8; 4 genes each assigned on SSC1, SSC7, SSC9, and SSC15; 3 genes each assigned on SSC2, SSC13 and SSCX; and 1 gene each assigned on SSC10, SSC11, and SSC16. Moreover, the present findings revealed 18 new chromosomal homologies between pig and human. Briefly, SSC3 regions were indicated to correspond with HSA1 and HSA10; SSC4 with HSA6; SSC5 with HSA2, HSA15, and HSA16; SSC6 with HSA3, HSA6, and HSA20; SSC7 with HSA11; SSC8 with HSA3, HSA6, and HSA7; SSC9 with HSA8; SSC13 with HSA1; SSC14 with HSA13; SSC15 with HSA19; SSC16 with HSA9.

Complete regression of xenografted human carcinomas by camptothecin analogue-carboxymethyl dextran conjugate (T-0128).

Clinically available camptothecins (CPTs), such as irinotecan (CPT-11) and topotecan, represent one of the most promising classes of antitumor agents, despite their toxicity. To improve their pharmacological profiles, a new macromolecular prodrug, denoted T-0128, was synthesized. This prodrug is a novel CPT analogue (T-2513)-carboxymethyl (CM) dextran conjugate via a triglycine spacer, with a molecular weight of Mr 130,000. This study was designed to test the concept that the rational design of a CPT-polymer conjugate would increase the efficacy of the parent drug. The in vivo antitumor study against Walker-256 carcinoma demonstrated that T-0128 was 10 times as active as T-2513, supporting this concept. Additionally, comparative efficacy studies of T-0128, T-2513, CPT-11, and topotecan were performed using a panel of human tumor xenografts in nude mice, showing the advantage of T-0128 over these CPTs. The maximal tolerated doses (MTDs) of T-0128, T-2513, and CPT-11 were comparable. Even a single i.v. injection of T-0128 at 6 mg/kg (based on the amount of T-2513 bound to CM dextran) induced complete regression of MX-1 mammary carcinoma. T-0128 at 10 mg/kg weekly for 3 weeks (one-tenth of its MTD) cured LX-1 lung carcinoma. Also, T-0128 below its MTD consistently cured or regressed St-4 gastric and HT-29 colorectal tumor xenografts that are highly refractory to CPTs. These demonstrate the broad range of therapeutic doses achieved with T-0128. Pharmacokinetic studies were performed to correlate the efficacy results obtained for T-0128 with plasma and tissue drug concentrations using Walker-256 tumor-bearing rats. Results showed that after i.v. administration of T-0128, the conjugate continued to circulate at a high concentration for an extended period, resulting in tumor accumulation. In the tumor, the sustained release of T-2513 occurred. In contrast, T-2513 disappeared rapidly from the body. The significant increases in the amount and exposure time of released T-2513 in the tumor explain well the enhanced efficacy of T-0128. In conclusion, this study indicated that T-0128 improved the potency of T-2513, demonstrating the proof of the above concept.